Hepatocarcinogenesis in vivo is now recognized to involve a complex, usually slowly developing sequence of linked cellular reactions that are entrained by a rapid initiating event and culminated by development of malignant lesions. Initiation is necessary, but not sufficient, to cause neoplastic transformation of hepatic epithelial cells. Separating the appearance of malignant tumors from the initiating event are a series of successive phenotypic alterations in the population of cells at risk that comprise the phases of tumor progression. Tumor progression may be viewed as a biological phenomenon essential to neoplastic transformation, during which initiated cells progressively develop phenotypic modifications that allow their selective multiplication. We have developed an in vitro transformation system, employing cultured hepatic epithelial cells, that reproduces a sequential process of tumor progression comparable to that in vivo. It is the goal of this proposed project to elucidate the mechanisms underlying the major cellular phenotypic variations arising during tumor progression by studying this process in transforming liver cells in vitro. Phenotypic variants studied will include a) increased resistance to the cytotoxic action of the carcinogen, b) more rapid and less regulated growth, and c) karyotypic instability. In addition the role of stem line selection and augmention in tumor progression will be analyzed.